Exposure of tyrosine residues in proteins. III. Reaction of cyanuric fluoride and N-acetylimidazole with ovalbumin, chymotrypsinogen, and trypsinogen

Gorbunoff, Marina J.

doi:10.1021/bi00834a050

Cited by 39 publications

(7 citation statements)

References 46 publications

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“…However, due to the fact that BTL2 is a multitryptophan (Trp = 9) protein and also rich in other aromatic amino acids (Phe = 16, Tyr = 18), assignment of the contributions of single Trp residues to near‐UV CD spectra might substantially be impeded by contributions of other aromatic residues . Nevertheless, strong transitions at 290 and 305 nm in the near‐UV region were previously identified for multitryptophan proteins such as chymotrypsin, and carboxypeptidase A . We similarly observed two distinct bands at 285 and 300 nm in the near‐UV region of BTL2 [Fig.…”

Section: Discussionsupporting

confidence: 53%

“…4 Despite the fact that zinc binding sites are rarely observed in lipases, 2 all of the identified members of the lipase family I.5 possess a zinc binding site. 5 As a result of having an extra domain for the zinc coordination, they exhibit larger molecular weights (40)(41)(42)(43)(44)(45) than other bacterial lipases (30 kDa). 2 Choi et al have studied this zinc binding site and clarified that the zinc coordinating residues along with the zinc ion itself have a profound effect on structural stability of the thermoalkalophilic lipase from Bacillus stearothermophilus (L1) at high temperatures.…”

Section: Introductionmentioning

confidence: 99%

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Probing the roles of two tryptophans surrounding the unique zinc coordination site in lipase family I.5

et al. 2015

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A unique zinc domain found in all of the identified members of the lipase family I.5 is surrounded by two conserved tryptophans (W61 and W212). In this study, we investigated the role of these hydrophobic residues in thermostability and thermoactivity of the lipase from Bacillus thermocatenulatus (BTL2) taken as the representative of the family. Circular dichroism spectroscopy revealed that the secondary structure of BTL2 is conserved by the tryptophan mutations (W61A, W212A, and W61A/W212A), and that W61 is located in a more rigid and less solvent exposed region than is W212. Thermal denaturation and optimal activity analyses pointed out that zinc induces thermostability and thermoactivity of BTL2, in which both tryptophans W61 and W212 play contributing roles. Molecular explanations describing the roles of these tryptophans were pursued by X-ray crystallography of the open form of the W61A mutant and molecular dynamics simulations which highlighted a critical function for W212 in zinc binding to the coordination site. This study reflects the potential use of hydrophobic amino acids in vicinity of metal coordination sites in lipase biocatalysts design.

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Section: Discussionsupporting

confidence: 53%

Section: Introductionmentioning

confidence: 99%

Probing the roles of two tryptophans surrounding the unique zinc coordination site in lipase family I.5

et al. 2015

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“…The reaction of CyF with proteins is carried out in aqueous buffered solution containing 10% dioxane to keep the reagent soluble, at pH values between 9 and 12 [3,4,9]. However this particular reagent does not lend itself well to an examination of Con A, the paramount difficulty being the high pH (minimally 9) at which the reaction is carried out.…”

Section: Reaction With Cyanuric Fluoride (Cyf)mentioning

confidence: 99%

“…lt was felt that the resulting CD spectra should then lend themselves to a more detailed view as to the origin of the various bands noted in native Con A. This approach has been used with considerable success in studies by Gorbunoff of several common protein systems [3,4].…”

Section: Introductionmentioning

confidence: 99%

Circular dichroism studies on chemically modified derivatives of concanavalin A

1972

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“…In spite of this, information about its secondary and tertiary structures was quantitatively limited till recent time. Several studies on the conformational changes in ovalbumin induced by certain denaturing agents have been reported (Gorbunoff 1969;Koseki et al, 1990;Batra et al, 1989b).…”

Section: Introductionmentioning

confidence: 99%

Relationship between functional properties and structure of ovalbumin

et al. 1994

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The effects of ovalbumin (OVA) denaturation using urea, guanidinium chloride (GdnHCl), sodium dodecyl sulphate (SDS), cetylpyridinium chloride (CPC), 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS), and 5 different cationic detergents with various side chains, HCl, and CH3COOH were observed. Progressive unfolding in ovalbumin was measured as a function of fluorescent light intensity, peak response and shift in the maximum of emission. Kinetic measurements demonstrated that the rate of denaturation usually followed a double exponential decay pattern, but at small concentrations of urea and acids first-order reaction was indicated. The reversibility of the unfolding-folding transitions was confirmed from tryptophan fluorescence and circular dichroism (CD) measurements. Differences in secondary structure were observed and changes of alpha-helical content were calculated. Polyacrylamide gel electrophoresis (PAGE) with and without sodium dodecyl sulphate (SDS-PAGE) showed differences in the structure of native and denatured ovalbumin. Native protein samples in PAGE demonstrated smaller number and larger mobilities of subunits than denatured ones with different reductants, such as SDS and 2-mercaptoethanol (2 ME). Scanning of SDS protein patterns showed the appearance of aggregated forms in region of 45 kD.

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Exposure of tyrosine residues in proteins. III. Reaction of cyanuric fluoride and N-acetylimidazole with ovalbumin, chymotrypsinogen, and trypsinogen

Cited by 39 publications

References 46 publications

Probing the roles of two tryptophans surrounding the unique zinc coordination site in lipase family I.5

Probing the roles of two tryptophans surrounding the unique zinc coordination site in lipase family I.5

Circular dichroism studies on chemically modified derivatives of concanavalin A

Relationship between functional properties and structure of ovalbumin

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